Journal: Disease Markers

Article Title: Evaluation of Immune Infiltration Based on Image Plus Helps Predict the Prognosis of Stage III Gastric Cancer Patients with Significantly Different Outcomes in Northeastern China

doi: 10.1155/2022/2893336

Figure Lengend Snippet: Expression of CD4, CD8, CD20, CD56, CD68, CD117, and CD177 by immune cells infiltrated tumor tissues of GC patients. Representative images of immune markers staining in immune cells from GC samples are shown at ×200 (100 mm) original magnification. Positive expression of CD4 (a), CD8 (b), CD20 (c), CD56 (d), CD68 (e), CD117 (f), and CD177 (g) infiltrated in tumor tissues with a regular distribution. In order to show the distribution from immune cells more clearly, we used computerized imaging system Image-Pro Plus version 6.2 to highlight positive areas in different colors.

Article Snippet: After cleaned in distilled water, the paraffin sections were pretreated with citrate buffer, pH 6.0 (CD177) and EDTA Antigen Retrieval Solution, pH 8.0 (CD4, CD8, CD20, CD56, CD68, and CD117) for 3 min at 120°C in a pressure cooker, and endogenous peroxidase was inhibited with 3% H 2 O 2 in PBS for 10 min. Nonspecific actions in the sections were also blocked with goat serum (BOSTER, USA) for 1 h at room temperature.

Techniques: Expressing, Staining, Imaging

Journal: Disease Markers

Article Title: Evaluation of Immune Infiltration Based on Image Plus Helps Predict the Prognosis of Stage III Gastric Cancer Patients with Significantly Different Outcomes in Northeastern China

doi: 10.1155/2022/2893336

Figure Lengend Snippet: The schematic diagram of the distribution of immune cells. The blue points represent CD20 + B cells, the yellow points represent CD4 + T cells, the orange points represent CD117 + mast cells, the green points represent CD68 + macrophages, the pink points represent CD8 + T cells, the black points represent CD56 + NK cells, and the red points represent CD177 + neutrophils. Although not all immune cells are distributed according to this fixed law, this pattern can basically reflect the general characteristics of their distribution.

Article Snippet: After cleaned in distilled water, the paraffin sections were pretreated with citrate buffer, pH 6.0 (CD177) and EDTA Antigen Retrieval Solution, pH 8.0 (CD4, CD8, CD20, CD56, CD68, and CD117) for 3 min at 120°C in a pressure cooker, and endogenous peroxidase was inhibited with 3% H 2 O 2 in PBS for 10 min. Nonspecific actions in the sections were also blocked with goat serum (BOSTER, USA) for 1 h at room temperature.

Techniques:

Journal: Disease Markers

Article Title: Evaluation of Immune Infiltration Based on Image Plus Helps Predict the Prognosis of Stage III Gastric Cancer Patients with Significantly Different Outcomes in Northeastern China

doi: 10.1155/2022/2893336

Figure Lengend Snippet: Differences in immune marker positive area/total area between the two groups (survival time of patients in group A was less than 1 year and group B survival time was more than 5 years) by the rank sum test. (a) CD4 + T cells ( P < 0.001). (b) CD8 + T cells ( P < 0.001). (c) CD20 + B cells ( P < 0.001). (d) CD68 + macrophages ( P < 0.001). (e) CD117 + mast cells ( P < 0.001). (f) CD177 + neutrophils ( P < 0.001).

Article Snippet: After cleaned in distilled water, the paraffin sections were pretreated with citrate buffer, pH 6.0 (CD177) and EDTA Antigen Retrieval Solution, pH 8.0 (CD4, CD8, CD20, CD56, CD68, and CD117) for 3 min at 120°C in a pressure cooker, and endogenous peroxidase was inhibited with 3% H 2 O 2 in PBS for 10 min. Nonspecific actions in the sections were also blocked with goat serum (BOSTER, USA) for 1 h at room temperature.

Techniques: Marker

Journal: Molecular Therapy

Article Title: VEGF Blockade Enables Oncolytic Cancer Virotherapy in Part by Modulating Intratumoral Myeloid Cells

doi: 10.1038/mt.2013.39

Figure Lengend Snippet: Analysis of rRp450 efficacy in sarcoma models. (a) Tumor cells were infected with rRp450 at the indicated MOI and harvested for HSV titer as determined by standard plaque assay at 1, 24, 48, and 72 hours post-infection (n = 4). Error bars represent SEM. (b) A673 cells were infected at the indicated MOI and cell viability was measured on days 2, 4, and 6 by MTT assay (n = 4). Error bars represent SD. (c) Mice bearing A673 tumors received two intratumoral injections of rRp450 at 1 × 107 pfu or PBS control, on days 0 and 2, and then followed for tumor growth (n = 7). (d) Mice bearing 143.98.2 tumors received two intratumoral injections of rRp450 at 1 × 107 pfu or PBS as a control on days 0 and 2, and were followed for tumor growth (n = 5–10). CR, complete response; ITu, intratumoral; MOI, multiplicity of infection; oHSV, oncolytic herpes simplex virus; PBS, phosphate-buffered saline; pfu, plaque-forming unit; PD, progressive disease; PR, partial response; SD, stable disease.

Article Snippet: Six to eight weeks old female athymic nude (nu/nu) mice (Harlan Sprague Dawley, Indianapolis, IN) were injected subcutaneously with A673 Ewing sarcoma cells (5 × 10 6 ) in 30% matrigel (BD Biosciences, San Diego, CA).

Techniques: Infection, Plaque Assay, MTT Assay

Journal: Molecular Therapy

Article Title: VEGF Blockade Enables Oncolytic Cancer Virotherapy in Part by Modulating Intratumoral Myeloid Cells

doi: 10.1038/mt.2013.39

Figure Lengend Snippet: Recruitment of myeloid cells in oHSV-injected tumors. A673 xenograft tumors were injected with rRp450 or PBS control. (a) The relative numbers of CD11b+ myeloid cells in the spleen (n = 4–5, t-test) and infected flank tumors (n = 6, t-test) were determined 3 days after virus injection by flow cytometry. In a separate experiment, (b,c) spleens and (d,e) tumors were collected 24 hours after virus injection and analyzed by hematoxylin and eosin (H&E) and by Ly6G immunohistochemistry staining (4X objective). The spleen served as a control, and brown staining is restricted to the red pulp and absent from the white pulp. oHSV, oncolytic herpes simplex virus; PBS, phosphate-­buffered saline.

Article Snippet: Six to eight weeks old female athymic nude (nu/nu) mice (Harlan Sprague Dawley, Indianapolis, IN) were injected subcutaneously with A673 Ewing sarcoma cells (5 × 10 6 ) in 30% matrigel (BD Biosciences, San Diego, CA).

Techniques: Injection, Infection, Flow Cytometry, Immunohistochemistry, Staining

Journal: Molecular Therapy

Article Title: VEGF Blockade Enables Oncolytic Cancer Virotherapy in Part by Modulating Intratumoral Myeloid Cells

doi: 10.1038/mt.2013.39

Figure Lengend Snippet: Induction of mVEGF in oHSV-injected tumors. A673 xenograft tumors were injected with rRp450 or PBS control and tumors were harvested at 3 days. The amount of (a) tumor-derived hVEGF (n = 7) and (b) stroma-derived mVEGF was determined by ELISA (n = 7). Error bars represent SEM. ELISA, enzyme-linked immunosorbent assay; hVEGF, human vascular endothelial growth factor; mVEGF, mouse vascular endothelial growth factor; oHSV, oncolytic herpes simplex virus; PBS, phosphate-buffered saline.

Article Snippet: Six to eight weeks old female athymic nude (nu/nu) mice (Harlan Sprague Dawley, Indianapolis, IN) were injected subcutaneously with A673 Ewing sarcoma cells (5 × 10 6 ) in 30% matrigel (BD Biosciences, San Diego, CA).

Techniques: Injection, Derivative Assay, Enzyme-linked Immunosorbent Assay

Journal: Molecular Therapy

Article Title: VEGF Blockade Enables Oncolytic Cancer Virotherapy in Part by Modulating Intratumoral Myeloid Cells

doi: 10.1038/mt.2013.39

Figure Lengend Snippet: Depletion of CD11b+ cells in oHSV-injected tumors. (a) A673 tumor-bearing animals were administered intraperitoneally either αβ-gal antibody (control) or α-CD11b antibody on the indicated days and either intratumoral PBS (control) or rRp450 (oHSV) on day 0. (b) Tumors were collected at day +3 and analyzed by flow cytometry. Shown are representative scatter plots from an n = 3 experiment illustrating the baseline CD11b+ and GR1+ populations, their depletion by anti-CD11b antibody injection, and their increase with oHSV infection. Only one scatter plot is shown for CD11b depletions because all six were essentially identical, with absence of CD11b cells regardless of PBS or oHSV injection. (c) Tumors were analyzed by ELISA for murine VEGF production (n = 5–8). Error bars represent SEM. ELISA, enzyme-linked immunosorbent assay; IP, intraperitoneal; IT, intratumoral; oHSV, oncolytic herpes simplex virus; PBS, phosphate-buffered saline; VEGF, vascular endothelial growth factor.

Article Snippet: Six to eight weeks old female athymic nude (nu/nu) mice (Harlan Sprague Dawley, Indianapolis, IN) were injected subcutaneously with A673 Ewing sarcoma cells (5 × 10 6 ) in 30% matrigel (BD Biosciences, San Diego, CA).

Techniques: Injection, Flow Cytometry, Infection, Enzyme-linked Immunosorbent Assay

Journal: Molecular Therapy

Article Title: VEGF Blockade Enables Oncolytic Cancer Virotherapy in Part by Modulating Intratumoral Myeloid Cells

doi: 10.1038/mt.2013.39

Figure Lengend Snippet: Treatment of A673 xenografts with combination bevacizumab and oHSV. Mice bearing A673 xenografts were treated with ITu oHSV, IP bevacizumab or a combination of both and followed for (a) tumor growth (n = 7–8) and (b) survival (n = 7–8). Error bars represent SEM. (c) Intratumoral virus production was measured in a separate cohort. Tumors were harvested at times shown and measured for infectious virus particles by plaque assay. Bevacizumab reduced virus production in the tumors, likely due to the antiangiogenic effects causing tumor cell death and limiting virus spread and production (n = 6). Error bars represent SEM. Bev, bevacizumab; IP, intraperitoneal; ITu, intratumoral; oHSV, oncolytic herpes simplex virus; pfu, plaque-forming unit.

Article Snippet: Six to eight weeks old female athymic nude (nu/nu) mice (Harlan Sprague Dawley, Indianapolis, IN) were injected subcutaneously with A673 Ewing sarcoma cells (5 × 10 6 ) in 30% matrigel (BD Biosciences, San Diego, CA).

Techniques: Plaque Assay

Journal: Molecular Therapy

Article Title: VEGF Blockade Enables Oncolytic Cancer Virotherapy in Part by Modulating Intratumoral Myeloid Cells

doi: 10.1038/mt.2013.39

Figure Lengend Snippet: Effects of the combination of intratumoral oHSV and intraperitoneal bevacizumab on the tumor vasculature. A673 tumor-bearing animals were administered either ITu PBS and IP rat IgG control antibody, ITu rRp450 and IP rat IgG control antibody, ITu PBS and IP bevacizumab or ITu rRp450 and IP bevacizumab. Tumors were harvested 3 days post-infection and either analyzed by ELISA for (a) hVEGF (n = 4–5) and (b) mVEGF (n = 4–6), evaluated by immunohistochemistry for apoptosis by TUNEL staining and for (c) pericytes by α-smooth muscle actin (α-SMA) staining or (d) endothelial cells by CD31 staining. Error bars represent SD (in a,b). Tumors were also quantified for (e) vessel numbers (n = 6, 10 high power fields per tumor) and (f) microvessel density (total vessel area per high power field; n = 6, 10 high power fields per tumor). Error bars represent SEM. Data were compared with PBS and only those indicated by an asterisk reached statistical significance. Bev, bevacizumab; ELISA, enzyme-linked immunosorbent assay; hpf, high power field; hVEGF, human vascular endothelial growth factor; mVEGF, mouse vascular endothelial growth factor; IP, intraperitoneal; ITu, intratumoral; oHSV, oncolytic herpes simplex virus; PBS, phosphate-buffered saline; TUNEL, terminal deoxyribonucleotidyl transferase-mediated dUTP nick end labeling.

Article Snippet: Six to eight weeks old female athymic nude (nu/nu) mice (Harlan Sprague Dawley, Indianapolis, IN) were injected subcutaneously with A673 Ewing sarcoma cells (5 × 10 6 ) in 30% matrigel (BD Biosciences, San Diego, CA).

Techniques: Infection, Enzyme-linked Immunosorbent Assay, Immunohistochemistry, TUNEL Assay, Staining, End Labeling

Journal: Molecular Therapy

Article Title: VEGF Blockade Enables Oncolytic Cancer Virotherapy in Part by Modulating Intratumoral Myeloid Cells

doi: 10.1038/mt.2013.39

Figure Lengend Snippet: Model of enhancement of oHSV efficacy by VEGF blockade. A model consistent with our data is that virus infection of tumor cells stimulates an innate neutrophilic infiltration and, through an unknown mechanism, depletes M1-type tumor-associated macrophages. Infection also induces production of host-derived mVEGF, either directly or indirectly via myeloid cells. The increase in stroma-derived mVEGF is an example of the virus effects on the local tumor microenvironment, but in the A673 model is overshadowed by tumor-derived hVEGF and likely plays only a minor role in angiogenesis. (We postulate that the effect of stroma-derived mVEGF may be more impactful in models with less tumor-derived hVEGF production). The virus-induced CD11b+ infiltrate results in production of protumor growth and proangiogenic factors from neutrophils and M2-type macrophages (+ factors represented by small arrows) that are no longer offset by M1-type macrophages. The combination of oHSV and anti-VEGF is more antiangiogenic, and the resulting tumor cell death is likely responsible for decreased intratumoral virus spread and production. Despite lower virus production, the combination results in improved antitumor effects due in part to modulation of the intratumoral myeloid cell composition, specifically by a mitigation of the decrease in tumoricidal M1-type macrophages. These effects are dependent on VEGFR2 signaling as they were seen with bevacizumab and r84. Modulation of myeloid cells is in part responsible for the combined effects of oHSV with VEGF blockade as it could be recapitulated by combining oHSV with depletion of CD11b cells, possibly by preventing a virus-induced predominance of M2-type compared with M1-type macrophages. The "?" denotes unknown mechanism. Bev, bevacizumab; hVEGF, human vascular endothelial growth factor; mVEGF, mouse vascular endothelial growth factor; oHSV, oncolytic herpes simplex virus; VEGFR, vascular endothelial growth ­factor receptor.

Article Snippet: Six to eight weeks old female athymic nude (nu/nu) mice (Harlan Sprague Dawley, Indianapolis, IN) were injected subcutaneously with A673 Ewing sarcoma cells (5 × 10 6 ) in 30% matrigel (BD Biosciences, San Diego, CA).

Techniques: Infection, Derivative Assay